In recent years, efforts have increasingly been made to produce industrially complex carbohydrates, such as secreted oligosaccharides. This has been due to the roles of such compounds in numerous biological processes in living organisms. Secreted oligosaccharides, such as human milk oligosaccharides (“HMOs”), have become particularly important commercial targets for nutrition and therapeutic applications. Human milk oligosaccharides (HMOs) have become of great interest in the past few years due to their important functions in human development. To date, the structures of at least 115 HMOs have been determined, and considerably more are probably present in human milk (Urashirna at al.: Milk oligosaccharides, Nova Science, 2011). Fucosylated lactoses including DFL are considered to be among the more important HMOs because of their nutritional value (see WO 2012/158517).
To date, ways of making large volumes of fucosylated lactoses at low cost have not been available. The isolation of fucosylated oligosaccharides from human milk has been rather difficult, even in milligram quantities, and very expensive due to the presence of a large number of other similar oligosaccharides in human milk. This problem has not been solved by current biotechnology or synthetic chemistry technology. Because of the growing commercial interest in nutritional compositions and supplements containing HMOs, there has been a need for a low cost method of making such HMOs.
Crystallization or recrystallization is one of the simplest and cheapest methods to separate a chemical product from contaminants and obtain it in pure form. In addition, crystalline modifications of a solid compound are important in the development of a compound, because different crystalline forms or polymorphs can have different properties—for example its thermodynamic stability, solubility, density, and hygroscopicity.
DFL was first isolated from mother's milk and its structure was elucidated with standard chemical methods including acid hydrolysis (Kuhn et al, Liebigs Ann, Chem. 611, 249 (1958)). Chemical synthesis of DFL has been disclosed by Takeo et al. (Carbohydr. Res. 141, 159 (1985)) and Fernandez-Mayoralas et al. (Carbohydr. Res. 154, 93 (1986)). The biosynthesis of DFL, together with 2′-FL and 3-FL (3-O-fucosyllactose), has been reported recently in NO 2012/112777. However, no crystalline form of DFL has, as yet, been described (Collins (ed.): Dictionary of Carbohydrates, Chapman & Hall/CRC (2006), p 675).
There has been a need, therefore, for a method of crystallizing DFL, particularly from a mixture of DFL and 2′-FL.